Automated EEG discontinuity in cooled newborns predicts cerebral MRI and neurodevelopmental outcome

Transcription

1 Automated EEG discontinuity in cooled newborns predicts cerebral MRI and neurodevelopmental outcome Jonathan M Dunne 1, David Wertheim MA, PhD 2 ; Paul Clarke MD, FRCPCH 3 ; Olga Kapellou MRCPCH, MD, PhD 4 ; Philippa Chisholm MRCPCH 4 ; James P Boardman PhD, FRCPCH 5 and Divyen K Shah MBChB, PhD 1,6 1 Barts and the London Medical School, London, UK 2 Kingston University, London, UK 3 Norfolk and Norwich University Hospitals NHS Foundation Trust, Norwich, UK 4 Homerton University Hospital, London, UK 5 MRC Centre for Reproductive Health, University of Edinburgh, UK 6 Royal London Hospital, London, UK Corresponding Author: Dr Divyen K Shah, Royal London Hospital, London E1 1BB. T: E: divyen.shah@bartshealth.nhs.uk Short title: EEG discontinuity in cooled newborns Key words: EEG, discontinuity, MRI, neurodevelopment, therapeutic hypothermia Abbreviations: TH therapeutic hypothermia Financial disclosure statement: The authors have no financial relationships relevant to this article to disclose. Funding source: JPB received funding support from Theirworld Potential conflicts of interest: None What is known about this topic: EEG discontinuity is associated with adverse neurologic outcomes in term infants after hypoxic-ischaemic encephalopathy. What this study adds: In term infants receiving therapeutic hypothermia, EEG discontinuity calculated using a novel algorithm at 24 hours is associated with cerebral tissue injury on MRI and with adverse neurodevelopmental outcomes. Therefore, it may provide a useful tool for early risk stratification when adjunctive therapies are most beneficial.

2 Contributors Statement: Jonathan M Dunne collected data, carried out the raw EEG data analysis, assisted with the statistical analysis, wrote the first draft of the manuscript and approved the final version of the manuscript. David Wertheim developed the EEG software used to measure discontinuity, assisted with writing the manuscript and approved of the final version of the manuscript. Paul Clarke, obtained ethics approval, collected data, assisted with writing the manuscript and approved the final version of the manuscript. Olga Kapellou reviewed the MR images and approved of the final version of the manuscript. Philippa Chisholm carried out neurodevelopmental testing and approved the final version of the manuscript. James P Boardman reviewed the MR images, assisted with writing the first draft of the manuscript, assisted with statistical analysis and approved the final version of the manuscript. Divyen K Shah conceived the study, collected data, assisted with the EEG reviews, statistical analysis, writing the first draft of the manuscript, approved of the final version of the manuscript and is guarantor. Word Count: 2498

3 Abstract Background and Hypothesis: Prolonged EEG discontinuity has been associated with poor neurodevelopmental outcomes after perinatal asphyxia but its predictive value in the era of therapeutic hypothermia (TH) is unknown. Hypothesis: In infants undergoing TH for hypoxicischaemic encephalopathy (HIE) prolonged EEG discontinuity is associated with cerebral tissue injury on MRI and adverse neurodevelopmental outcome. Method: Retrospective study of term neonates from 3 UK centres who received TH for perinatal asphyxia, had continuous 2 channel aeeg with EEG for a minimum of 48 hours, brain MRI within 6 weeks of birth, and neurodevelopmental outcome data at a median age of 24 months. Mean discontinuity was calculated utilising a novel automated algorithm designed for analysis of the raw EEG signal. Results: Of 49 eligible infants, 17 (35%) had MR images predictive of death or severe neurodisability (unfavourable outcome) and 29 (59%) infants had electrographic seizures. In multivariable logistic regression, mean discontinuity at 24 hours and 48 hours (both p=0.01), and high seizure burden (p=0.05) were associated with severe cerebral tissue injury on MRI. A mean discontinuity >30s per minute-long epoch, had a specificity and positive predictive value of 100%, sensitivity of 71% and a negative predictive value of 88% for unfavourable neurodevelopmental outcome at a 10µV threshold. Conclusions: In addition to seizure burden, excessive EEG discontinuity is associated with increased cerebral tissue injury on MRI and is predictive of abnormal neurodevelopmental outcome in infants treated with TH. The high positive predictive value of EEG discontinuity at 24 hours may be valuable in selecting newborns with HIE for adjunctive treatments.

4 Introduction Hypoxic-ischaemic encephalopathy (HIE) affects up to 6 per 1000 live births in industrialised nations and is an important cause of disability in survivors 1 2. Mild therapeutic hypothermia (TH) is a safe and effective neuroprotective intervention and is now standard care in resourcerich settings 3 4. TH increases the number of survivors without neurodisability, with the benefits persisting into childhood 5. The number needed to treat is nine 4, so there is a need to stratify patients who may stand to benefit from adjunctive therapies, and to improve the precision of prognostic information for families and clinicians. In the precooling era, prolonged EEG discontinuity of 30s or longer in term newborns with HIE 6, as well as in other conditions including cerebral haemorrhage and infection 7, was associated with unfavourable neurodevelopmental outcomes. However, the applicability of EEG quantification for infants undergoing TH after HIE is unclear. Continuous limited channel EEG with amplitude-integrated EEG (aeeg) monitoring is routinely used at the bedside in newborns undergoing TH for assessment of electrocortical background and to screen for seizures 8. We have previously shown that electrographic seizure burden is independently associated with cerebral tissue injury on MRI in term-born infants undergoing TH 8. Severely abnormal EEG and aeeg background patterns are associated with adverse outcome 9 10, but the precise features of the background that predict poor outcome are uncertain and qualitative assessments lack diagnostic precision and are susceptible to inter-observer variation. In order to improve the objectivity of EEG assessment in the newborn, we developed and applied computer software for automated quantification of EEG discontinuity. We aimed to test the hypothesis that an objective measure of EEG discontinuity is associated with cerebral tissue injury on MRI and neurodevelopmental outcome.

5 Method We studied infants who received TH between October 2007 and July 2011 in three tertiary UK Neonatal Intensive Care Units (NICUs): the Royal London Hospital, Homerton University Hospital, and Norfolk and Norwich University Hospital. Data were obtained as part of standard clinical care. This study had Research Ethics Committee approval (UK REC reference 14/EE/0205). Subjects Neonates were eligible for inclusion if they were born at 36 weeks gestation, treated with whole body TH for moderate to severe HIE (defined using standard criteria) 11, had aeeg monitoring within 12 hours of birth, and had brain MRI within 6 weeks. Exclusion criteria included death prior to MR imaging, images degraded by motion artefact, major congenital anomaly or a primary diagnosis of an inborn error of metabolism. The cohort is a subset of a larger cohort reported previously 8. Infants received intensive care and TH according to local guidelines, which were informed by the UK TOBY Cooling Register Clinician s Handbook 12. Sentinel events were defined as a sustained (pre-terminal) fetal bradycardia necessitating delivery of the infant, antepartum haemorrhage secondary to placental abruption, placenta praevia, cord prolapse or rupture, uterine rupture and shoulder dystocia. aeeg/eeg monitoring and discontinuity analysis Recordings obtained from the aeeg monitors (BRM2/3 BrainZ Instruments, Natus Medical Inc., CA, USA) were reviewed off-line (by DS and JD) to visually identify seizure-free and minimum artefact 2-hour epochs at 24 and 48 hours of age. Single channel cross-cerebral (P3- P4) EEG data were exported to Excel, and continuity was analysed in 1-minute epochs using software that we developed using MATLAB (The MathWorks, Inc., MA, USA), similar to that validated in previous work by comparing with visual assessment 13. The system detected an

6 interval if the absolute amplitude of the EEG was less than 15 µv with respect to the baseline for at least 6 seconds and therefore discontinuous; the analysis was repeated with a 10 µv threshold. The 10 and 15 µv thresholds were chosen to reflect the fact that the EEG from healthy term newborns is represented by more lower voltage high frequency wave forms in contrast to preterm or encephalopathic infants 14. For each recording the mean of the total interval length per epoch, the discontinuity value, was calculated and expressed in seconds. For an interval to be detected the minimum period of 6 seconds was chosen in order to avoid including normal short quiescent periods seen in tracé alternant 15. In order to allow assessment of temporal variability in EEG discontinuity, the length of the analysis epochs was one minute; intervals that started in the preceding epoch and continued into a subsequent period were included in the analysis. EEG discontinuity greater than 30 s per minute was regarded as a predominantly discontinuous trace, as this equates to 50% discontinuity within a minute implying that the EEG is predominantly discontinuous at the given amplitude threshold. This is similar to approaches previously applied in visual assessment of term infant EEG recordings 7. For four traces, the analysis was repeated which yielded the same mean interval duration. MR Imaging Magnetic resonance imaging was performed at local centres with conventional T1-weighted and T2-weighted sequences acquired at 1.5 Tesla. Anonymised images were rated independently by two investigators (OK and JPB), who were blind to the clinical data. The pattern of MRI injury was classified into two groups, using the system described by Rutherford et al. 16, which has prognostic value in the era of TH. This method includes rating the posterior limb of the internal capsule (PLIC), basal ganglia and thalami (BGT) and the subcortical white matter. Group 1 had a severe pattern of injury that predicts poor outcome, defined by the classification system as death or one or more of: mental development index (MDI) score less than 70 ( 2 SD below the mean) on the Bayley Scales of Infant Development (BSID II); score of 3 5 on the gross motor function classification system (GMFCS); or bilateral cortical visual

7 impairment with no useful vision. Group 2 had either normal images or less severe patterns of injury associated with normal or mildly abnormal (favourable) neurodevelopmental outcome. These patterns of injury were used to assign infants into either the favourable or unfavourable MR images outcome group. Neurodevelopmental Outcomes For the purposes of neurodevelopmental outcomes, infants were classified in the unfavourable outcome group if they had cerebral palsy that impaired independent walking; severe seizure disorder; hearing impairment requiring hearing aids or bilateral cortical visual impairment with no useful vision. Infants with better outcomes were placed in the favourable outcome group. Neurodevelopmental outcomes were assessed at multiple local centres, with tools of assessment including both BSID and Griffiths. Statistical Analysis Data were analysed using SPSS V22 (IBM). We investigated discontinuity at 10 and 15 µv thresholds at both 24 and 48 hour time epochs, seizure burden, Apgar score at 10 minutes, use of anticonvulsants, worst arterial cord or infant ph and base deficit within the first hour in univariable analyses with cerebral tissues injury on MRI using χ 2 statistic for categorical variables and analysis of variance (ANOVA) for continuous variables. Seizures were classified as previously reported for this cohort 8. Seizures were defined as rhythmic spike and/or wave activity on the raw EEG lasting at least 10 s in the absence of artefact. A low seizure burden was defined as no seizures or sporadic seizures lasting < 15 minutes in a single hour. A seizure episode lasting 15 minutes per hour for any hour during the period of monitoring including the cooling and rewarming period was classified as high seizure burden. Significant factors from the univariable analyses were examined in a binary logistic regression model to calculate odds ratios (OR), assessing their association with MRI outcome. The area under the receiver operating characteristic (ROC) was calculated for the relationship between

8 discontinuity >30s and unfavourable MRI outcomes. The OR, positive predictive value (PPV), negative predictive value (NPV), sensitivity and specificity were calculated for the relationship between discontinuity >30s and unfavourable neurodevelopmental outcomes. Two-sided p values were regarded as significant at the 0.05 threshold.

9 Results Of 52 eligible infants, three were excluded due to incomplete EEG recordings at 48 hours. The clinical characteristics of the remaining 49 are described in Table 1 and the EEG discontinuity measures, MRI findings and neurodevelopmental outcomes are detailed in the Table 2. MRI Outcomes Seventeen (35%) of the study cohort had MR images predictive of unfavourable outcome. On univariable analysis, factors associated with the unfavourable pattern of MRI outcome were high seizure burden (p=0.003), discontinuity at 24 hours (p<0.001) and at 48 hours (p<0.001) at 15 µv threshold (Table 3). In multivariable logistic regression high seizure burden (OR 4.2, 95% CI ; p=0.05), mean discontinuity at 24 hours (OR 1.04, 95% CI ; p=0.01) and at 48 hours (OR 1.05, 95% CI ; p=0.01) were associated with severe cerebral tissue injury on MRI (using the method described by Rutherford et al 16 ). Of infants with unfavourable MRI outcomes, the median (IQR) discontinuity in seconds was 31.2 (0.9, 51.8) and 43.5 (8.6, 56.5) at 10 and 15 µv respectively at 24 hours, and 2.7 (0.1, 42.7) and 20.3 (1.9, 53.2) at 48 hours (Figure 1). A mean discontinuity >30 s per 1-minute epoch had a PPV of 90% and 86% at 24 and 48 hours respectively (10 µv threshold) and of 75% and 80% at 24 and 48 hours (15 µv threshold) for group 1 (unfavourable MRI outcome), The area under the ROC curve at both thresholds and both time points was between 0.78 and 0.81 showing a high predictive value for discontinuity at both time points and thresholds. To investigate the possibility of collinearity between seizure burden and discontinuity (24 hours, 10 µv threshold) in the regression model we calculated correlation between these two variables and found the Pearson coefficient to be low, 0.30 (p=0.036).

10 Neurodevelopmental Outcomes Neurodevelopmental outcomes were available for 43/49 (88%) of the children (Table 2). Median age (interquartile range) of last follow-up was 24 (24, 24) months. 14/43 (33%) children had unfavourable clinical outcomes: four had died, six had cerebral palsy, two required bilateral hearing aids for profound sensori-neural hearing loss, one had autism, seizures and global speech delay and one had seizures and unilateral hearing loss (Table 2). Ten of the 14 children with unfavourable outcomes had a discontinuous trace (>30s). For these 14 children, the median (range) discontinuity in seconds was 50.3 (9.6, 55.1) and 55.2 (24.8, 58.0) at 10 and 15 µv respectively at 24 hours and 18.1 (0.1, 47.7) and 42.8 (5.3, 53.3) at 48 hours (Figure 2). A mean discontinuity >30 s per 1-minute epoch has a PPV of 100% and 87% at 24 and 48 hours respectively (10 µv threshold) and of 91% and 90% at 24 and 48 hours (15 µv threshold) for unfavourable neurodevelopmental outcome (Table 3). Of the 14 children with unfavourable clinical outcomes, 10 had MR images predictive of unfavourable outcome. Of the remaining four children, two had profound bilateral hearing loss and one had seizures and unilateral hearing loss, which the MRI system does not predict. One infant died at 6 months from complications of evolving cerebral palsy. For the 29 children with favourable neurodevelopmental outcomes only one had EEG discontinuity >30s at both time epochs and voltage thresholds. Six of these 29 had MRIs that would predict likely neurodisability: three of these children had neurodevelopmental abnormalities including mildly impaired cognition, and/or delayed speech and language development, and three were classified as normal at the time of assessment. Overall, the correlation between MRI and neurodevelopmental outcome was good: Pearson s R=0.62 (p<0.001). Of the six children lost to follow-up, five had MRI predictive of favourable outcomes.

11 Discussion Our study indicates that excessive EEG discontinuity assessed objectively with our automated software is associated with increased cerebral tissue injury on MRI, and is independently predictive of abnormal neurodevelopmental outcome in infants with HIE who have undergone TH. This association was statistically significant at 24 and 48 hours of age, at 10 V and 15 V thresholds. Using a discontinuity cut-off of 30 s, PPVs of 75-90% were obtained for MRIinferred outcomes and even higher PPVs for neurodevelopmental outcomes. The sensitivity for adverse neurodevelopmental outcome was greatest at 24 hours at 71% whereas the specificity at this time was at least 97%. This highlights the importance of early EEG monitoring in identifying infants who are likely to benefit from therapeutic interventions. Among infants with unfavourable MRI outcomes, the median discontinuity values dropped from 31.2 and 43.5 seconds at 24 hours to 2.7 and 20.3 s at 48 hours at 10 V and 15 V respectively, i.e. to below 30 s at both voltage thresholds. Hence in infants undergoing TH, the PPV of a discontinuous trace for cerebral tissue injury on MRI is greater at 24 hours than at 48 hours with a substantial number of babies with poor MRI outcomes having EEG discontinuity values less than 30 s at 48 hours. This method may provide an important tool for risk stratification in early postnatal life when adjunctive therapies are most likely to be beneficial. In earlier work we found that high seizure burden (seizures longer than 15 minutes per hour) was associated with brain tissue injury 8, but we were unable to investigate the contribution of discontinuity, and its co-occurrence with seizure burden, to tissue injury and outcome due to technical limitations. The novel automated software we have developed for reliable quantification of discontinuity from cotside recordings of the EEG has now enabled such analysis: the evidence for collinearity between seizure burden and discontinuity was weak, which suggests that the point estimates of effect of each of these predictor variables in the model are likely to be reliable. Our novel automated software analysed single channel cross-

12 cerebral EEG data, therefore it could be utilised with other aeeg/eeg software from which the raw EEG data can be extracted. Previous studies have shown a relationship between features of multichannel EEG background 17 as well as limited channel EEG/aEEG measures 18 and MRI outcomes in neonatal encephalopathy in the pre-cooling era, in that increased EEG depression is associated with more severe abnormality on cerebral MRI. Studies have shown the predictive value of EEG discontinuity lasting at least 30 s with adverse neurodevelopmental outcomes in infants with HIE 6 as well as in groups of infants with broader neurologic diagnoses 7 in the precooling era. Menache et al. 7 reported that all 10 babies (100%) with a predominant interburst interval duration of 30 s or greater had adverse neurologic outcomes; however, in that study the analogue EEG recordings may have been shorter with a minimum duration of 30 minutes. The use of automated quantification of background EEG activity has previously been demonstrated in infants In a qualitative multichannel video-eeg study, Nash et al. 9 found that a persistent burst suppression or continuous low voltage pattern beyond 24 hours of age in infants undergoing TH was highly predictive of brain injury on MRI. Using qualitative aeeg background patterns, a summary trend of the raw EEG, for term infants who had undergone TH, Thoresen et al. 10 showed that the PPV of an abnormal aeeg background for poor outcome in cooled infants was >90% at 24 hours in contrast to normothermic infants in whom the PPV of an abnormal trace reached 90% by age 12 hours. In the present study using an unbiased objective measure of discontinuity, we found similar PPVs at 24 and 48 hours of age in cooled babies and the PPV of the lower voltage threshold was greater. Although our study was primarily designed to investigate the relationship between EEG discontinuity and MRI outcomes, we were also able to show an independent significant relationship between discontinuity and neurodevelopmental outcomes with a high PPV at 24 hours. We also found high concordance between MRI outcomes and later clinical outcomes at neurodevelopmental follow-up. The high early PPV of EEG discontinuity for abnormal MRI

13 and neurodevelopmental outcomes may be a useful tool for early stratification of asphyxiated newborn babies for potentially beneficial adjunctive neuroprotective therapies. Acknowledgements We are grateful to Dr Courtney Wusthoff for her assistance with rating the seizure burden. We are grateful to all the nursing and medical staff from the study sites and all the families.

14 References 1. Thornberg E, Thiringer K, Odeback A, et al. Birth asphyxia: incidence, clinical course and outcome in a swedish population. Acta Paediatrica 1995;84(8): Levene ML, Kornberg J, Williams THC. The incidence and severity of post-asphyxial encephalopathy in full-term infants. Early Human Development 1985;11(1): Garfinkle J, Sant'Anna GM, Wintermark P, et al. Cooling in the real world: Therapeutic hypothermia in hypoxic-ischemic encephalopathy. European Journal of Paediatric Neurology 2013;17(5): Edwards AD, Brocklehurst P, Gunn AJ, et al. Neurological outcomes at 18 months of age after moderate hypothermia for perinatal hypoxic ischaemic encephalopathy: synthesis and metaanalysis of trial data. British Medical Journal 2010; Azzopardi D, Strohm B, Marlow N, et al. Effects of Hypothermia for Perinatal Asphyxia on Childhood Outcomes. New England Journal of Medicine 2014;371(2): Wertheim D, Mercuri E, Faundez JC, et al. Prognostic value of continuous electroencephalographic recording in full-term infants with hypoxic-ischemic encephalopathy. Archives of Disease in Childhood 1994;71(2):F97-F Menache CC, Bourgeois BFD, Volpe JJ. Prognostic value of neonatal discontinuous EEG. Pediatric Neurology 2002;27(2): Shah DK, Wusthoff CJ, Clarke P, et al. Electrographic seizures are associated with brain injury in newborns undergoing therapeutic hypothermia. Archives of Disease in Childhood-Fetal and Neonatal Edition 2014;99(3):F219-F Nash KB, Bonifacio SL, Glass HC, et al. Video-EEG monitoring in newborns with hypoxic-ischemic encephalopathy treated with hypothermia. Neurology 2011;76(6): Thoresen M, Hellstrom-Westas L, Liu X, et al. Effect of Hypothermia on Amplitude-Integrated Electroencephalogram in Infants With Asphyxia. Pediatrics 2010;126(1):E131-E Azzopardi DV, Strohm B, Edwards AD, et al. Moderate hypothermia to treat perinatal asphyxial encephalopathy. The New England journal of medicine 2009;361(14): UK TOBY Cooling Register Clinician s Handbook pdf. 13. Wertheim DFP, Eaton DGM, Oozeer RC, et al. A new system for cotside display and analysis of the preterm neonatal electroencephalogram. Developmental Medicine and Child Neurology 1991;33(12): Werner S, Bickford R, Stockard-Pope J. Atlas of Neonatal Electroencephalography. New York: Raven Press, Andre M, Lamblin MD, d'allest AM, et al. Electroencephalography in premature and full-term infants. Developmental features and glossary. Neurophysiologie Clinique-Clinical Neurophysiology 2010;40(2): Rutherford MA, Pennock JM, Counsell SJ, et al. Abnormal magnetic resonance signal in the internal capsule predicts poor neurodevelopmental outcome in infants with hypoxic-ischemic encephalopathy. Pediatrics 1998;102(2): Biagioni E, Mercuri E, Rutherford M, et al. Combined use of electroencephalogram and magnetic resonance imaging in full-term neonates with acute encephalopathy. Pediatrics 2001;107(3): Shah DK, Lavery S, Doyle LW, et al. Use of 2-channel bedside electroencephalogram monitoring in term-born encephalopathic infants related to cerebral injury defined by magnetic resonance imaging. Pediatrics 2006;118(1): Lofhede J, Thordstein M, Lofgren N, et al. Automatic classification of background EEG activity in healthy and sick neonates. Journal of Neural Engineering 2010;7(1).

15 Table 1: Characteristics of study infants Perinatal Characteristics (total neonates = 49) Male sex 32 (65%) Median birth weight, grams (range) 3360 ( ) Median Apgar score at 10 min (IQR), n=44 6 (4, 8) Median first ph, by cord or within 1h, mmol/l (IQR), n=48 Median first base deficit, by cord or within 1h, mmol/l (IQR), n= (6.81, 7.10) (-11.3, -21.5) Need for respiratory support at 10min age 37 (76%) Median age when heart rate first >100/min, minutes (IQR), n=48 2 (1, 5) Median age at first gasp, minutes (range) 5 (0, 20) n=41 Antenatal sentinel event identified 8 (16%) Suspected clinical seizures 36 (73%) Treated with anticonvulsants 37 (76%) Received sedation during cooling 42/46 (91%) Median age at start of aeeg monitoring (range), n=47 4h 50m (15m to 11h 20 m) Median duration of aeeg monitoring (range), n=48 89h 53m (13h 20m to 168h 58m) Any seizures identified on aeeg/eeg 29 (59%) High seizure burden on aeeg/eeg 18 (37%) Epoch analysed at 24 and 48 hours, minutes 119 (80, 120) (range) Median age at MRI, days (interquartile range) 11 (8, 14) Severe pattern of injury on MRI (group 1) 17/49 (35%) Unfavourable neurodevelopmental outcome 14/43 (33%)

16 Table 2: Discontinuity values in seconds, MRI, seizure burden and neurodevelopment for the study babies. Baby DC 10 V at 24h DC 15 V at 24h BGT PLIC White Matter Seizure Burden Neurodevelopmental Outcome Mild Normal Mild None Normal Mild Normal Moderate None Normal Mild Normal Moderate None Normal Mild Normal Normal None Normal Mild Normal Mild None Normal Mild Normal Normal Low Normal Normal Normal Mild Low Normal Mild Normal Mild None Normal Mild Normal Moderate None Bilateral SN hearing loss Normal Normal Normal None n/a Normal Normal Normal None n/a Normal Normal Normal None Normal Mild Normal Normal None Normal Normal Normal Severe High Normal Mild Normal Moderate High Normal Mild Normal Moderate Low Seizures and unilateral hearing loss Normal Normal Mild None Normal Moderate Partially Impaired Severe High Normal Mild Normal Moderate High Normal Moderate Normal Severe High Mild speech delay Mild Normal Normal None Normal Normal Normal Normal None Normal Mild Normal Mild None n/a Mild Normal Mild High Normal Normal Partially Impaired Normal None Normal Moderate Normal Mild None Normal Normal Normal Mild Low Normal Severe Severely Impaired Severe High Dyskinetic CP Normal Normal Severe High Normal Moderate Normal Moderate None n/a Normal Normal Normal Low Bilateral SN hearing loss, significant receptive and expressive speech delay Mild Normal Moderate None Normal Mild Normal Mild High Normal Normal Normal Normal Low n/a Mild Normal Mild High Mild delay Mild Normal Mild Low Normal Moderate Severely Impaired Mild None Normal Normal Normal Mild Low n/a Mild Normal Mild High Normal Severe Severely Impaired Moderate Low Quadriplegic CP Moderate Severely Impaired Mild High Spastic quadriplegic CP, GDD, SN hearing loss and dystonia Severe Severely Impaired Moderate High GDD, microcephaly, visual impairment, dystonia with epilepsy and GORD Severe Severely Impaired Moderate High Quadriplegic CP Severe Severely Impaired Severe Low Died Moderate Severely Impaired Severe High Global speech delay, autism and seizures Moderate Severely Impaired Moderate High Died Severe Severely Impaired Moderate Low Died Mild Partially Impaired Normal High Died Severe Severely Impaired Moderate High Spastic quadriplegic CP

17 DC: Discontinuity (at stated threshold and time epoch). Low seizure burden includes either no or sporadic seizures, high seizure burden includes either frequent seizures or status epilepticus. BGT: Basal ganglia and thalami, PLIC: Posterior limb of the internal capsule, CP: Cerebral palsy, GDD: Global developmental delay, GORD: Gastro-oesophageal reflux disease. SN, sensori-neural.

18 Table 3: Univariable analyses for the association between risk factors and poor outcome on MRI (first grid) and multivariable regression model assessing significant factors in univariable analyses against poor MRI outcome (second grid). Predictive values for unfavourable outcome with a mean discontinuity >30s on MRI (third grid) and neurodevelopmental outcome (fourth grid). Univariable Analysis Independent χ 2 or ANOVA* Statistic P Value Seizure Burden Apgar 10 *F= Lowest cord or infant ph in first hour *F= Worst base deficit in first hour *F= Use of anticonvulsants Binary Logistic Regression OR Lower CI Upper CI P Value Seizure Burden μv at 24 hours μv at 24 hours μv at 48 hours μv at 48 hours Predictive Values of Discontinuity (>30s) for Unfavourable MRI Outcomes Sensitivity Specificity PPV NPV Area under ROC P Value 10 μv at 24 hours 53% 94% 82% 79% μv at 24 hours 53% 91% 75% 78% μv at 48 hours 41% 97% 87% 76% 0.78 < μv at 48 hours 47% 94% 80% 77% Predictive Values of Discontinuity (>30s) for Unfavourable Neurodevelopmental Outcomes Sensitivity Specificity PPV NPV OR 95%CI 10 μv at 24 hours 71% 100% 100% 88% μv at 24 hours 71% 97% 91% 88% μv at 48 hours 50% 97% 87% 80% μv at 48 hours 64% 96% 90% 85% , , , OR: Odds ratio, CI: Confidence interval, PPV: Positive predictive value, NPV: Negative predictive value, ROC: Receiver operating characteristic.

19 Figure 1: Plots comparing mean discontinuity and MRI outcome (favourable or unfavourable). The left panels shows mean discontinuity at 24 hours and the right panels at 48 hours. The top panels utilise a 10 V threshold and the bottom panels a 15 V threshold. The box edges show the interquartile range and the whiskers the range.

20 Figure 2: Plots comparing mean discontinuity and neurodevelopmental outcome. The left panels shows mean discontinuity at 24 hours and the right panels at 48 hours. The top panels utilise a 10 V threshold and the bottom panels a 15 V threshold. The box edges show the interquartile range and the whiskers the range.

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